PROJECT SUMMARY The central goal of this project is to investigate the mechanisms of T regulatory cells (Tregs) and interleukin 35 (IL-35) in attenuating lung inflammation and remodeling leading to WHO type-2 pulmonary hypertension (PH), and arresting the transitional process from left ventricular (LV) failure to right ventricular (RV) hypertrophy and failure. While most studies in the chronic heart failure (CHF) field focus on left ventricle, we recently demonstrated that LV failure causes profound lung inflammation, fibrosis, and severe type-2 PH. Remarkably, in some cases lung remodeling is so extensive that the lung tissue becomes as solid as liver tissue in CHF animals. We have now obtained solid evidence that Tregs are effective in suppressing LV failure-induced lung inflammation, and type-2 PH in animals with existing LV failure, but the mechanisms remain unknown. IL-35 is a heterodimer composed of IL-12? (p35) and Epstein-Barr virus-induced gene-3 (EBI3) subunits. IL-35 can effectively promote the formation of Tregs and regulatory B cells (Bregs). Therefore, IL-35 may play an important role in attenuating lung inflammation/remodeling, type-2 PH, and the transition from LV failure to RV hypertrophy and failure through both Treg-dependent or Treg-independent mechanisms. Here we will investigate how Tregs and IL-35 affect CHF-induced lung inflammation, and type- 2 PH. Specific Aim-1 will mainly determine the mechanisms of Tregs and IL-35 in suppressing lung inflammation, type-2 PH, and the transition from LV failure to RV failure in mice with existing LV failure. We will perform the following studies: (i) examine the impact of Treg depletion on IL-35 production, T cell and dendritic cell activation, lung inflammation, type-2 PH, and the transition from LV failure to RV hypertrophy and/or failure in mice, (ii) determine why the ratio of Tregs to T effector memory cells is dramatically reduced in lung tissue in CHF animals, and whether this can be explained by alterations in Treg proliferation or apoptosis, and (iii) determine the mechanism of IL-35 in suppressing lung remodeling and type-2 PH in mice with existing LV failure with a focus on Tregs and T cell activation. Specific Aim-2 will determine the crosstalk between Tregs and IL-35, and their corresponding roles in regulating lung inflammation and type-2 PH in mice with existing LV failure. We will perform following studies: (i) Determine if IL-35 is required for the optimal protection by Treg induction in attenuating lung inflammation, type-2 PH, and the transition from LV failure to RV hypertrophy/failure. (ii) determine the role of IL-35-independent Tregs in attenuating lung inflammation, type-2 PH and CHF progression, and (iii) determine whether administration of IL-35 is sufficient to rescue Treg depletion-induced lung inflammation and type-2 PH in mice with existing LV failure. Successful completion of this project will provide exciting and novel insights into cardiovascular protective mechanism(s) of Tregs and IL-35, and demonstrate the therapeutic potential of targeting Tregs and IL-35 to halt or reverse lung inflammation, and the transition from LV failure to RV hypertrophy and/or failure.